Proteomic Changes in Paracoccidioides brasiliensis Induced by Fludioxonil

Paracoccidioides brasiliensis is a pathogenic fungus capable of thermodimorphic transition, causing systemic mycoses in humans. Fludioxonil, a phenylpyrrole fungicide, inhibits this transition and affects genes related to fungal cell wall composition. This study explored the proteomic response of P. brasiliensis yeast cells exposed to fludioxonil, identifying over 4100 proteins, approximately 50% of the total proteome, through DIA (data-independent acquisition) mass spectrometry. Treatment led to significant proteomic changes, with 28 proteins upregulated and 26 downregulated. Upregulated proteins were linked to oxidative and osmotic stress responses, including the mitochondrial stress response. HSP12, the most upregulated protein, participates in osmotic stress adaptation via the HOG1 pathway, while CsbD and Memo 1 family proteins were associated with stress response and fungal invasiveness. Fludioxonil exposure increased reactive oxygen species (ROS) production and catalase activity, suggesting oxidative stress adaptation. Additionally, mitochondrial membrane hyperpolarization was observed, indicating mitochondrial dysfunction. The treatment also triggered cell wall remodeling, with reduced levels of mannosyltransferase, β-glucosidase, and Chitinase, enzymes involved in carbohydrate metabolism, while chitin synthase levels were increased. These findings reveal that fludioxonil disrupts both mitochondrial metabolism and cell wall integrity, shedding light on its antifungal mechanism in P. brasiliensis.